Pluto Bioinformatics

GSE110380: Proximity-CLIP provides a snapshot of occupied cis-acting elements on RNA in different subcellular compartments on a transcriptome-wide scale

Bulk RNA sequencing

Many cellular RNAs localize to specific subcellular compartments. Currently, methods to systematically study subcellular RNA localization are limited and lagging behind proteomic approaches. Here, we combined APEX2-mediated proximity biotinylation of proteins with PAR-CLIP to simultaneously profile the proteome and the transcriptome bound by RNA binding proteins in any given subcellular compartment. Our approach is fractionation-independent and does not rely on additional RNA manipulation and labeling steps, thus making it easy to apply. Furthermore, it enables to study the locali-zation of RNA processing intermediates, as well as the identification of regulatory RNA cis-acting elements occupied in different cellular compartments. In a proof-of-concept study we studied RNA and protein localization in the nucleus, cytoplasm and at cell-cell interfaces using Proximity-CLIP. These experiments revealed among other in-sights frequent transcriptional readthrough continuing for several kilobases down-stream of the canonical cleavage and polyadenylation site, a differential binding pat-tern of nuclear and cytoplasmic mRNAs, as well as the localization of mRNAs contain-ing 3UTR CUG sequence elements at cell-cell interfaces, of which many encode regulatory proteins. SOURCE: Markus Hafner (markus.hafner@nih.gov) - Laboratory of Muscle Stem Cells and Gene Regulation National Institutes of Health